I have designed a few pairs of sgRNA oligos using both CHOPCHOP and CRISPOR. I checked for their alignments within themselves. However, when I did the Primer-BLAST using both oligos (excluding the flanking sticky sequences). Only 1 of the 4 oligo pairs showed the gene of interest without any mismatch. All other pairs showed mismatch with non-specific genes, ironically these were ranked in the top in the sgRNA tool result. What may be the reason for mismatches and non-specific alignment? Please suggest.

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